The long-term goal is to study the role of nitric oxide as a mediator of vascular processes and intercellular communication. In order to study these effects in intact animals, I plan to use homologous recombination to disrupt the NOS gene, generating mice that are genetically incapable of expressing the gene. I have cloned the mouse NOS gene and constructed a targeting vector containing a mutant NOS gene. This gene will be introduced into pluripotent mouse embryonic stem cells, which will be microinjected into blastocysts and implanted into foster mother mice. Germ-line transmission of the disrupted NOS gene from chimeric mice will result in heterozygous offspring, which may be bred to obtain mice homozygous for the defect, NOS (-) mice. I plan to examine selected features of the development and function of the vascular system and the nervous system in these NOS (-) mice: histologic analysis of vascular and nervous system morphogenesis, and hemodynamic studies in intact animals. This project builds upon my past research experience in molecular biology and my clinical training and interest in cardiology. However, it also involves the fields of developmental biology, mouse genetics, and vascular biology, all very different from my previous background. This experience will complement my past research background and greatly enrich my research training.